Abstract
Indiscriminate collection of Arnebia benthamii roots from wild for extraction of red pigments by traders prompted to put this Himalayan plant species under critically endangered category. Concerning the development of sustainable phytoconstituents sources, the present study is aimed at understanding the factors crucial for the scalability of A. benthamii cell suspension cultures. Here, the effect of orbital shaking speed (60–80 rpm) and inoculum density (5–15%) on leaf-induced cell suspension cultures during sequential scale up in shake flask (0.25–5 L) was assessed. The suspension cultures were established on Murashige and Skoog medium supplemented with 10 μM 6-benzylaminopurine and 5 μM indole-3-butyric acid having a 20-days cultivation cycle. The data revealed a significantly (p ≤ 0.05) high biomass productivity (523.80 ± 5.95 g/L, fresh weight) in 0.25 L vessel with 10% inoculum at 70 rpm as compared to large (5.0 L) volume flask (310.80 ± 32.85 g/L). Interestingly, the optimum inoculum density was found to vary with the size of culture vessels. Furthermore, noticeable effects of sequential scaleup with shaking speed and inoculum density were also observed on hydrogen peroxide (stress indicator) production as well as enzymatic (catalase and phenylalanine ammonia-lyase activity) and non-enzymatic (phenolic compounds) antioxidant potential. A. benthamii cells cultivated with optimized conditions had a significant amount of red pigment, with 669.17 ± 26.71 mg dry weight/L yield of total shikonin derivatives. In conclusion, the results clearly demonstrated the likelihood of scalability using optimized process variables for the production of natural red pigments using A. benthamii cell suspension cultures.
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Abbreviations
- CAT:
-
Catalase
- DPPH:
-
2,2-Diphenyl-1-picrylhydrazyl
- DW:
-
Dry weight
- H2O2 :
-
Hydrogen peroxide
- MS:
-
Murashige and Skoog
- PAL:
-
Phenylalanine ammonia-lyase
- RGR:
-
Relative growth rate
- ROS:
-
Reactive oxygen species
- TPC:
-
Total polyphenol contents
- APM:
-
Arnebia pigment production medium
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Acknowledgements
Authors are greatly acknowledging the guidance and support received from Director, CSIR-IHBT. RK and JD are also thankful to CSIR for Ph.D. fellowship.
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The study was financially supported by Council of Scientific and Industrial Research (CSIR), Government of India vide project No. MLP-0144 and MLP 201.
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RK executed the experiments, collected data and written the manuscript. JD participated in data interpretation and revising of the manuscript. DK has performed the UPLC analysis. SB made substantial contributions from conception and design of this study to data interpretation and reviewing the manuscript.
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Kumar, R., Devi, J., Kumar, D. et al. Arnebia benthamii cell suspension cultures as a source of natural red pigments: optimization of shaking speed and inoculum density to maximize process productivity during sequential scaleup. Plant Biotechnol Rep 17, 353–367 (2023). https://doi.org/10.1007/s11816-023-00835-8
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DOI: https://doi.org/10.1007/s11816-023-00835-8